Material and methods

A total number of 565 barley accessions were selected for screening with P. neglectus, comprising of 375 winter, 30 spring barley lines (H. vulgare ssp. vulgare) and 160 wild species accessions (H. vulgare ssp. spontaneum). As a result of limited glasshouse facilities, it was impossible to test all accessions at a time. The plant material was grouped to its geographical origin (four groups): European barley accessions (218), North American barley accessions (129), Asian-African barley accessions (213) and Australian barley accessions (5). As a result of the large number of plants, the 565 accessions had to be tested in three different experiments. Each experiment contained a representative number of accessions from each geographical region. For each experiment, about 186 to 190 accessions were tested in a completely randomized block design. The accessions of each experiment were tested as single plant in six replications. A validation experiment was carried out in the following way: 5% of the most resistant accessions of each experiment, regarding their overall rank, were selected and tested with six repeats under the conditions described above. All the experiments were conducted in the glasshouse and in the climate chamber with 23°C day and 18°C night temperature. Seeds were germinated on wet filter paper at 26°C for 1 d in the dark. Each seedling was placed in a 20 cm³ tube (12 cm (H) height × 2 cm (∅) thickness 2.3 mm) filled with steam-sterilized sand. Seedlings were inoculated with 400 P. neglectus juveniles in 1 ml water medium after 10 d. Further steps were followed as described in Keil et al. (Reference Keil, Laubach, Sharma and Jung2009). After 12 weeks, the plants were uprooted and nematodes were extracted from the sand and the roots were chopped using a Baermann funnel placed in a misting chamber for 5 d. Nematode suspension were collected in a bottle and placed at 5°C before counting. For QTL mapping, 126 F₁ anther-derived DH lines from a cross Igri × Franka were used (Graner et al., Reference Graner, Jahoor, Schondelmaier, Siedler, Pillen, Fischbeck, Wenzel and Herrmann1991). A molecular map was constructed by integrating diversity array technology (DarT) markers into the already available Igri × Franka map (Stein et al., Reference Stein, Prasad, Uwe, Thiel, Zhang, Wolf, Kota, Varshney, Perovic, Grosse and Graner2007). QTL analysis was carried out by composite interval mapping using the program QTL Cartographer V2.5 (Wang et al., Reference Wang, Basten and Zeng2010).

Results and discussion

A representative collection of 565 cultivated and wild barley accessions was tested for P. neglectus resistance in three different experiments. The mean number of nematodes/plant in the three experiments was 3335, 2920 and 1546 (P = 0.0001), respectively. Fifty per cent of the accessions had 1564 to 2988, 5% had below 982 and another 5% had above 5048 nematodes/plant. There was no barley accession without any nematode infection, i.e. showing immunity to P. neglectus. The number of nematodes/plant ranged from 350 to 12,000. In a verification experiment with 32 barley accessions, five accessions namely ‘BCB-39’, ‘AC Queens’, ‘BYDV 17’, ‘AC Legend’ and ‘Beysehir’ were identified as moderately resistant. Among the ten least susceptible accessions, three Turkish and only one German accessions were found (Keil et al., Reference Keil, Laubach, Sharma and Jung2009). In general, German accessions had a tendency for high susceptibility to P. neglectus infection, reflecting the lack of selection pressure. In Turkey, breeders have selected for resistance towards Pratylenchus species over decades due to a high infection pressure. On the other hand, in Germany, nematode resistance was not a breeding aim because this problem arose only recently mainly due to narrow crop rotations.

To unravel the genetics of RLN resistance, an Igri × Franka DH population was tested under greenhouse and climate chamber conditions. The means of nematode counts of both parents, Igri (861) and Franka (1591), were significantly different (P < 0.05). Among the DHs, a high phenotypic variation was observed for P. neglectus infection. Transgressive segregants were also observed in the population which indicates that favourable alleles are dispersed between both parental lines. For mapping, DArT markers and previously mapped restriction fragment length polymorphisms were used. Five QTLs were mapped with a likelihood of odds score between 2.71 and 6.35 and explaining phenotypic variation of 8 to 16%, as shown in the Fig. 1(a–d). Some QTL positions are coincident with previously identified QTLs or major genes conferring resistance to a diverse spectrum of barley pathogens such as scald (Rhynchosporium secalis, Rrs) (Graner and Tekauz, Reference Graner and Tekauz1996; Garvin et al., Reference Garvin, Brown, Raman and Read2000), Pyrenophora teres (net type blotch disease; Graner and Tekauz, Reference Graner and Tekauz1996), and barley yellow dwarf virus (Ryd2; Collins et al. Reference Collins, Paltridge, Ford and Symons1996). The data provide clear evidence of a polygenic inheritance of RLN resistance in barley with major QTL having a big impact on infection rates. The tightly linked markers flanking the QTLs will be turned into diagnostic markers for marker-assisted selection of resistant plants from segregating offspring.

Fig. 1 (a–d) Chromosomal location of QTL for P. neglectus resistance in the barley DH population Igri × Franka.